A. actinomycetemcomitans, a gram-negative bacterium, is the causative agent of localized aggressive periodontitis (LAP). A. actinomycetemcomitans is also a member of the important HACECK group of bacteria implicated in enfective endocarditis. One of its virulence factors, a potent leukotoxin, kills specifically white blood cells and can also destroy erythrocytes. Leukotoxin is a member of the RTX toxins. Much of the present knowledge in the biosynthesis of leukotoxin in A. actinomycetemcomitans is based on similarity with other bacterial systems, such as E. coli, but some differences exist between those systems and A. actinomycetemcomitans. We propose to study the biosynthesis of leukotoxin in A. actinomycetemcomitans to expand our knowledge of the physiology and pathogenesis of this organism which can lead to a better understanding of the role played by leukotoxin as virulence factor. More specifically, we propose studies that aim to: 1. Characterize of novel genes involved in leukotoxin biosynthersis. We have isolated leukotoxin mutants, and several mutations map somewhere outside of the leukotoxin operon. We propose experiments that will identify and characterize the nature of the mutations. 2. Determine the function of ItxC in the modification and activiy of leukotoxin. We propose experiments that will allow us to study modification of leukotoxin in ItxC mutants and to determine the role of ItxC on the activity of leukotoxin. This study will expand our knowledge of the genetics of leukotoxin production in A. actinomycetemcomitans. A. actinomycetemcomitans, an oral pathogen, causes periodontal and heart disease. We are studying the genetics involved in the synthesis of a toxin produced by this bacterium. It is our goal to understand the role played by this toxin in the disease with the hopes of enhancing our ability to prevent or treat diseases caused by A. actinomycetemcomitans.